The invention relates to novel enzyme inhibiting compounds, their synthesis, and their use in treating pathological and physiological conditions.
Pyrimidine analogs and pyrimidine nucleosides are widely used as chemotherapeutic agents for cancer and for viral, fungal, bacterial and parasitic infections. Most pyrimidine analogs used in cancer chemotherapy must be convened to the nucleoside 5xe2x80x2-monophosphate level before any anticancer activity can be realized. However, all most all are administered as nucleosides or bases to facilitate transport into cells. The administered compounds are subject to catabolism and inactivation by various enzymes within a patient""s body. Pyrimidines, for example, are degraded by the enzymes uridine phosphorylase (UrdPase) and dihydrouracil dehydrogenase (DHUDase). As a result, the balance between the anabolic (activation) and catabolic (inactivation) pathways must be considered when designing or choosing a chemotherapeutic regime for treating various malignancies, or for treating viral, fungal, bacterial or parasitic infections.
Until recently, most studies of pyrimidine analog metabolism have focused on anabolism, with little attention devoted to catabolism. Pyrimidine bases and nucleoside analogs can be anabolized within a patient""s body to the nucleoside 5xe2x80x2-monophosphate, or catabolized to xcex2-amino acids. The catabolism of nucleosides to bases proceeds by nucleoside phosphonilases. The resulting bases are then convened to their respective xcex2-amino acids by a chain of three reactions, catalyzed by DHUDase, dihydropyrimidinase and xcex2-ureidopropionase. Wastemack, Pharmac. Ther., 8:629-651 (1981); Naguib, et al, Cancer Res., 45:5405-5412 (1985). Cytidine, cytosine and their analogs must be deaminated before they can be catabolized.
The importance of DHUDase as a target Ibr chemotherapy has been established by several recent studies. For example, patients receiving continuous infusion of 5-fluorouracil (5-FUra) at a constant rate were found to have plasma concentrations of 5-FUra that varied significantly during treatment. This variation followed a circadian rhythm which was the inverse of that observed for DHUDase activity. Harris et al, Biochem. Pharmac., 37: 759-4762 (1988); Harris et al, Cancer Res., 49:6610-6614 (1989); Petit E., et al Cancer Res., 48:1676-1679 (1988); Naguib et al, Biochem. Pharmac., 45: 667-673. (1993). That is, high plasma concentration of 5-FUra was associated with low DHUDase activity and vice versa. A significant correlation between the circadian rhythm of DHUDase activity and that of the anticancer efficacy of 5-FUra and 5-fluoro-2xe2x80x2-deoxyuridine (5-FdUrd) has also been reported. Petit et al. Cancer Res., 48:1676-1679 (1988); von Roemeling et al, Advances in Chronobiology, Part B, 357-373 (1987). Thus it is clear that a strong association exists between the level of DHUDase activity and the bioavailability and efficacy of fluoropyrimidines for chemotherapy.
The importance of DHUDase in cancer chemotherapy is further emphasized by studies with inhibitors of DHUDase, where the inhibitors were found to increase the concentration and half life of 5-FUra in plasma and to dramatically enhance the chemotherapeutic efficacy of 5-FUra in vitro and in vivo. Nevertheless, coadministration of known inhibitors of DHUDase with 5-FUra has not been popular due to several drawbacks associated with such previously known inhibitors. Although the known inhibits enhanced the antitumor activity of 5-FUra, they also served as alternate substrates and caused substantial host-toxicity. Cooper et al, Cancer Res., 32:390-397 (1972); Gentry et al, Cancer Res., 31:909-912 (1971). It was also believed that DHUDase inhibition would mimic the genetic deficiency of this enzyme which is known to be accompanied by neurological disorders. Bakkeren et al, Clinica Chimica Acta, 140:246-247 (1984); Tuchman et al N. Engl. J. Med, 313:245-249 (1985); Diasio et al, J. Clin. Invest., 81:47-51 (1988); Wadman et al, Adv. Exp. Med. Biol., 165A: 109-114 (1984). Finally, it was generally believed that tumors lack or possess very little DHUDase activity. Chaudhury et al, Cancer Res., 18:318-328 (1958); Heidelberger et al, Cancer, Res., 30:1549-1569 (1970); Mukherjee et al, Biol. Chem., 235:433-437 (1960).
Thus, despite the potential promise of DHUDase inhibitors for chemotherapy regimes, currently known inhibitors have demonstrated several drawbacks that have discouraged their use in such treatments.
UrdPase inhibitors are also known to possess a number of clinically useful attributes. For example, UrdPase Inhibitors have been proposed to increase the selectivity and efficacy of various uracil and uridine derivatives in cancer chemotherapy. U.S. Pat. No. 5,077,280 (Sommadossi et al) discloses that UrdPase inhibitors can be used as rescue agents to reduce the toxicity of antiviral agents such as 3xe2x80x2-azido-3xe2x80x2-deoxythymidine (AZT), Ideal UrdPase inhibitors are those that are potent, specific, and non toxic. Moreover, useful UrdPase inhibitors should be readily soluble in aqueous solutions buffered within the physiological pH range.
As noted above halogenated pyrimidine bases such as 5-FUra and halogenated pyrimidine nucleosides such as 5-FdUrd have been used as chemotherapeutic agents in cancer treatments. Because these compounds are subject to rapid degradation, efficacy of the compound is reduced. Also, the catabolites of these chemotherapeutic agents (e.g., 2-fluoro-xcex2-alanine) are believed to be more toxic to a patient""s healthy cells.
Halogenated pyrimidine nucleosides, for example, are known to share the same catabolic pathway as uridine. Because there is little functional thymidine phosphorylase in many tumor cells, the first step in the catabolic pathway in tumor cells relies primarily on UrdPase. The inhibition of this enzyme in tumor cells serves to inhibit the catabolism of the chemotherapeutic agents in tumor tissue, thereby increasing their effectiveness. In healthy host tissue, the halogenated pyrimidine nucleosides can still be catabolized to their pyrimidine counterparts by the action of thymidine phosphorylase.
Similarly, halogenated pyrimidine bases such as 5-FUra can compete with cellular pyrimidines and their nucleotides for incorporation into RNA and DNA. However, UrdPase inhibitors increase the plasma and uridine concentration (Monks et al, Biochem. Pharmac., 32, 2003-2009) (1983); Darnowski et al, Cancer Res., 45:5364-5368 (1985)) and the availability of uridine for salvage of host healthy tissue. The increase in plasma uridine concentration also increases the pool of uracil nucleosides in tissue. The increased intracellular uridine concentration can thus reduce the toxicity of halogenated compounds in host tissue. Moreover, it has been shown that the addition of a phosphorylase inhibitor selectively increases the ability of host tissue to salvage uridine. Darnowski et al, Cancer Res., 45:5364-5368 (1985). This tissue specific enhancement of uridine utilization is of particular importance for chemotherapy regimes using 5-fluorouracil.
Another application of UrdPase inhibitors lies in their use in the protection against host toxicity of various antiviral agents. For example, viral therapies for patients infected with the human immunodeficiency virus (HIV) and/or those suffering from Acquired Immune Deficiency Syndrome (AIDS) have typically involved the administration of an antiviral pyrimidine nucleoside such as AZT. Such an antiviral agent functions by inhibiting the reverse transcriptase enzyme of the HIV to reduce the cytopathic effects of the virus.
The utility of antiviral pyrimidine nucleosides such as AZT has been limited by the toxic effects of AZT or its catabolites such as 3xe2x80x2 -amino-3xe2x80x2-deoxythymidine (AMT) on uninfected cells. Cretton et al, Molec., Pharmac., 39:258-266 (1991). Prolonged administration of such compounds produces severe side effects including the suppression of bone marrow growth and severe anemia. The dosage and duration of AZT therapies is limited because of such complications.
It is now known that uridine and, to some extent, cytidine can reverse the toxic effects of AZT in human bone marrow progenitor cells without affecting the antiviral activity of AZT in infected cells. Sommadossi et al Antimicrob. Agents Chemother., 32, 997-1000 (1988). This rescuing effect of uridine, although generally beneficial, has disadvantages because of the body""s rapid uridine catabolism. Consequently, high doses are required, and high doses of uridine can cause serious toxic side effects, including phlebitis and pyrogenic reactions.
Viral therapies that combine AZT or similar compounds with UrdPase inhibitors have been suggested in U.S. Pat. No. 5,077,280 (Sommadossi et al). Such treatments utilize UrdPase inhibitors to maintain effective levels of uridine in plasma sufficient to rescue uninfected cells without requiring the administration of high, potentially harmful doses of uridine.
Further, a number of synthetic UrdPase inhibitors have been proposed. See Niedzwicki et al, Biochem Pharmac., 31:1857 (1982); Naguib et al, Biochem Pharmac., 36:2195 (1987); Naguib et al, Biochem. Pharmac., 46:1273-1283 (1993). U.S. Pat. No. 4,613,604 (Chu et al); and U.S. Pat. No. 5,141,943 (Naguib et al). Such UrdPase inhibitors include a variety of substituted acyclouridines and 5-benzyl barbiturate derivatives.
Substituted acyclouridines are good inhibitors of UrdPase, but tend to have limited water solubility and are difficult and expensive to synthesize. Water solubility is essential for practical chemotherapy and treatment of infection in order to enable intravenous administration at physiological pH ranges and to allow formulation of reasonable volumes to be administered. Unfortunately, some acyclouridines, such as benzyl acyclouridine and its derivatives, are soluble only to about 1 mM in water at room temperature. Administration of a physiologically useful dose would require dilution of these compounds into excessively large volumes. 5-Benzyl barbiturate derivatives are also useful UrdPase inhibitors and have been found to be more water soluble and more desirable than derivatives of benzyl acyclouridine.
The maintenance of or increase in plasma uridine levels is also useful to treat several pathological and physiological conditions. For example, uridine has been shown to increase myocardial performance, glucose uptake, glycogen synthesis and the breakdown of ATP in heart tissue of rabbits. Plasma uridine level fluctuations also have important implications in muscle performance and in myocardial contractility. Further, uridine levels are important in central nervous system functioning. For example, the control of intracellular and plasma uridine levels is believed to have important implications in the treatment of CNS disorders, including cerebrovascular disorder and convulsions, epilepsy, Parkinson""s and Alzheimer diseases, and senile dementias. Uridine is also potentially useful in the treatment of liver damage and hepatitis. (See Naguib et al, Biochem. Pharmac. 46:1273 (1993) and references cited therein).
It is thus apparent that it is desirable to inhibit the enzymes that rapidly degrade certain chemotherapeutic agents or that otherwise contribute to excess uracil or uridine catabolism. In particular, inhibitors of DHUDase and UrdPase are of great relevance to treatment regimes for cancers as well as viral, fungal, bacterial and parasitic infections. Further, the control of and maintenance of plasma uridine levels is thus important in treating and preventing many diseases and pathological conditions. UrdPase Inhibitors can also be used to increase available plasma uridine concentrations. As a result, there is a need for new and improved enzyme inhibiting compounds, particularly inhibitors of DHUDase and UrdPase.
Accordingly, it is an object of the invention to provide new compounds useful as DHUDase and UrdPase inhibitors. A further object of the invention is to provide such DHUDase and UrdPase inhibitors which can be used with various chemotherapy regimes to reduce the toxicity of chemotherapeutic agents to normal and uninfected cells. Another object of the invention is to provide methods for increasing the efficacy of chemotherapy regimes in treating cancers as well as viral, fungal, bacterial, and parasitic infections. A further object of the invention is to increase the efficacy of certain chemotherapeutic regimes while reducing adverse patient affects associated with such treatments. Yet another object of the invention is to provide methods for synthesizing such new inhibitors of DHUDase and UrdPase. It is also an object of the invention to provide methods and compositions useful to increase plasma uridine concentrations and effective useful to treat various physiological and pathological conditions. A further object of the invention is to provide methods to treat and/or prevent symptoms of inherited disorders of pyrimidine catabolism. These and other objects of the invention will be apparent from the description that follows.
The invention relates to novel compounds that are effective as inhibitors of DHUDase or UrdPase. The novel compounds are represented by the formula 
where X is S or Se, Y is H, I, F, Cl, Br, methoxy, benzyl, selenenylphenyl, or thiophenyl; and R1 is H or an a cyclo  acyclo tail having the general formula 
where R2 is H, CH2OH or CH2NH2; R3 is OH, NH2 or OCOCH2CH2CO2H; and R4 is O, S, or CH2.
Novel compounds of the invention that inhibit DHUDase include 5-(phenylselenenyl)uracil (PSU); 5-(phenylthio)uracil (PTU); 5-(phenylselenenyl)barbituric acid; and 5-(phenylthio)barbituric acid.
Preferred compounds of the invention that inhibit UrdPase include compounds of the above general formulas having an a cyclo tail. Such compounds include 1-[(2-hydroxyethoxy)methyl]-5-(phenylselenenyl)uracil (PSAU); 1-[(2-hydroxyethoxy)methyl]-5-(phenylthio)uracil (PTAU); 1-[(2-hydroxyethoxy)methyl]-5-(phenylselenenyl)barbituric acid; and 1-[(2-hydroxyethoxy)methyl]-5-(phenylthio)barbituric acid.
In another embodiment the invention relates to pharmaceutical compositions comprising a chemotherapeutic agent, such as a pyrimidine, in an amount effective to treat cancer or a viral, fungal, bacterial, or parasitic infection; an effective amount of a novel DHUDase or UrdPase inhibitor of the present invention; and a pharmaceutically acceptable carrier. The chemotherapeutic agent can be one that is commonly used to treat cancer or viral, fungal, bacterial or parasitic infections and which is subject to degradation within a patient""s body by DHUDase or UrdPase. Examples of such chemotherapeutic agents include pyrimidine compounds such as 3xe2x80x2-azido-3xe2x80x2-deoxythymidine; 3xe2x80x2-fluoro-3xe2x80x2-deoxythymidine; 2xe2x80x2, 3xe2x80x2-dideoxycytidin-2xe2x80x2-ene; 3xe2x80x2-deoxythymidin-2xe2x80x2-ene; 3xe2x80x2-azido-2xe2x80x2,3xe2x80x2-dideoxyuridine; 2xe2x80x2,3xe2x80x2-dideoxy-5-fluoro-3xe2x80x2-thiacytidine; 2xe2x80x2,3xe2x80x2-dideoxy-3xe2x80x2-thiacytidine; 5-fluoro-2xe2x80x2,3xe2x80x2-dideoxycytidine; 5-fluorouracil; 5-fluoro-2xe2x80x2-deoxyuridine; and heterodimers thereof and enantiomers thereof. The chemotherapeutic agent can also be a prodrug of pyrimidine nucleobase analogs, including 1,(2-tetrahydrofuryl)-5-fluorouracil (TEGAFUR); 5-fluorocytosine; 5xe2x80x2-deoxy-5-fluorouridine; and 1-ethoxymethyl-5-fluorouracil. The chemotherapeutic agent can also be a prodrug sold by Taiho Pharmaceutical Company, Ltd. of Osaka, Japan under the tradename UFT, which is a combination of 1,(2-tetrahydrofuryl)-5-fluorouracil and uracil.
In another embodiment the invention comprises a method for administering chemotherapeutic agents while protecting and/or rescuing normal or uninfected cells from any toxicity that may result from the administration of the chemotherapeutic agent. Further, methods are provided for improving the efficacy of the chemotherapeutic agent. The methods of the invention comprise administering the chemotherapeutic agent, and coadministering or sequentially administering a DHUDase or UrdPase inhibiting compound of the type disclosed herein. The inhibition of the activity of DHUDase or UrdPase prevents or slows the degradation of the chemotherapeutic agent by these enzymes. This prevents or slows the degradation of the chemotherapeutic agent also results in lower levels of potentially toxic catabolites of the chemotherapeutic agent. These methods thus facilitate a higher concentration and/or a longer half-life of the chemotherapeutic agent, thus increasing the efficacy of the treatment regime. An additional benefit is that any toxic side effects of the chemotherapeutic regime are minimized.
The use of enzyme inhibiting compounds of the present invention are also effective to provide increased plasma levels of natural pyrimidines, such as uridine, which can help to protect and/or rescue healthy cells from toxicity induced by chemotherapeutic agents. The administration of these compounds to increase plasma levels of natural pyrimidines can also be effective to treat pathological and physiological disorders that respond to the administration of such pyrimidines. Such disorders responsive to these treatments include CNS disorders, Parkinson""s disease, Alzheimer""s disease, senile dimentia, sleep disorders, muscle dysfunction, long disorders, diabetes, cardiac insufficiency and myocardial infarction, liver disease and liver damage.
In addition to the novel compounds disclosed herein it has also been discovered that several known compounds are effective as UrdPase inhibitors. The UrdPase inhibiting activity of such compounds was previously unknown.
The present invention also contemplates the synthesis of novel enzyme inhibiting compounds such as 5-(phenylselenenyl)uracil and 5-(phenylthio)uracil.